流体剪切力对MC3T3-E1细胞OPG、RANKL蛋白表达的实验研究

发布时间：2018-06-03 14:17

本文选题：流体剪切力 + 骨保护素　；参考：《兰州大学》2010年硕士论文

【摘要】： 目的：探索流体剪切力(Fluid Shear Stress, FSS)作用下,两种调节骨骼重建的重要分子骨保护素(osteoprotegerin, OPG)和破骨细胞分化因子(osteoclast differentiation factor, ODF)即细胞核因子kB受体活化因子配体(receptor activator of NF-kB ligand, RANKL)的蛋白表达情况,从而揭示流体剪切力对于维持骨组织形成与吸收动态平衡的特殊作用。方法：本研究采用体外模型对MC3T3-E1细胞加载流体剪切力,经过不同时长(Omin、30min、60min、90min、120min)加力后,对MC3T3-E1细胞分别进行了染色和裂解,然后运用免疫荧光和蛋白印迹法对骨保护素(OPG)和破骨细胞分化因子(RANKL)的蛋白表达水平进行定量分析。结果：静态培养组RANKL的表达随时间的延长而增加,OPG的表达则减少,使得OPG与RANKL的比值(OPG/RANKL)不断下降,促使骨组织向着加速吸收的方向发展；加载流体剪切力30min后,蛋白检测发现FSS促进了OPG蛋白表达且抑制了RANKL蛋白表达,阻滞了OPG/RANKL比值减少的趋势：加力60min后,发现FSS进一步促进了OPG的表达,同时RANKL的表达则受到较大抑制,与静态组相比此时OPG/RANKL的比值增加了50%左右,使得骨组织向着成骨的方向发展；在120min内,随着FSS作用时间的延长OPG/RANKL的比值也在不断的增加,各组间OPG与RANKL的比值(OPG/RANKL)有显著的提高(P0.05)。结论：成骨细胞对FSS的响应要经历一个短暂的潜伏期,但与其他作用力相比流体剪切力对成骨细胞的作用更加有效；流体剪切力作用使得OPG的表达增加,RANKL的表达减少,二者的协同作用(OPG/RANKL)在成骨细胞和破骨细胞联合调节骨骼形成和吸收的过程中扮演着重要角色,从而进一步说明FSS对OPG和RANKL的作用影响着骨组织形成与吸收的动态平衡。
[Abstract]:Objective: to explore the effect of fluid Shear Stress, FSS) on shear stress. The protein expression of osteoprotegerin (OPG) and osteoclast differentiation factor, ODF), the nuclear factor kB receptor activator ligand receptor of NF-kB ligand (RANKLL), are two important molecules regulating bone remodeling. Thus, the special role of fluid shear stress in maintaining the dynamic balance of bone formation and absorption is revealed. Methods: in this study, the MC3T3-E1 cells were stained and lysed with fluid shear force (FSF) loaded with the model in vitro, and the MC3T3-E1 cells were treated with the stress for 30 min, 90 min or 120 min respectively, and the MC3T3-E1 cells were stained and lysed respectively after the stress was applied to the MC3T3-E1 cells for 30 min, 90 min and 120 min, respectively. Then the protein expression levels of osteoprotegerin (OPG) and osteoclast differentiation factor (RANKL) were quantitatively analyzed by immunofluorescence and Western blot. Results: in the static culture group, the expression of RANKL increased and decreased with the increase of time, and the ratio of OPG to RANKL decreased, and the bone tissue developed towards the direction of accelerated absorption. Protein detection showed that FSS promoted the expression of OPG protein and inhibited the expression of RANKL protein, and blocked the decrease of OPG/RANKL ratio. After adding 60min, it was found that FSS further promoted the expression of OPG, while the expression of RANKL was inhibited greatly. Compared with the static group, the ratio of OPG/RANKL increased by about 50%, which made the bone tissue develop toward the direction of osteogenesis, and in 120min, the ratio of OPG to RANKL increased with the prolongation of the time of FSS, and the ratio of OPG to RANKL increased significantly (P 0.05). Conclusion: the response of osteoblasts to FSS has a short incubation period, but fluid shear stress is more effective than other forces on osteoblasts, and fluid shear stress can increase the expression of OPG and decrease the expression of RANKL. The synergistic action of OPG / RANKL plays an important role in the process of osteoblasts and osteoclasts jointly regulating bone formation and absorption, which further indicates that the effect of FSS on OPG and RANKL affects the dynamic balance of bone formation and absorption.
【学位授予单位】：兰州大学
【学位级别】：硕士
【学位授予年份】：2010
【分类号】：R329

【参考文献】